The participation of herpes simplex virus (HSV) glycoproteins in T cell-mediated lysis of HSV-infected syngeneic target cells was examined by using a temperature sensitive (ts) mutant defective in glycoprotein synthesis at the nonpermissive temperature (39 degrees C), and 2-deoxy-D-glucose and tunicamycin, known inhibitors of both HSV replication and glycoprotein synthesis in HSV-infected cells. Lymphocytes cytotoxic for HSV-infected cells lysed C57BL/6 Wt-3 cells infected with the mutant, ts A1, at 39 degrees C less efficiently than at 33 degrees C. Treatment of HSV-infected C57BL/6 Wt-3 cells with 1% 2-deoxy-D-glucose for 14 hr reduced their susceptibility to T cell-mediated lysis by 73% in the 51Cr release assay. Treatment of HSV-infected C57BL/6 Wt-3 cells with 0.2 microgram/ml tunicamycin for 14 hr reduced their susceptibility to T cell-mediated lysis by 78% in the 51Cr release assay. The reduction in T cell-mediated lysis by 2 deoxy-D-glucose and tunicamycin was found not to be due to the effects of the compounds on the H-2 antigens. We conclude that HSV specific glycoproteins are involved in T cell-mediated lysis of HSV-infected cells.